RNAseq from patients

NDUFB11 · 03-05-2019, 07:50 AM

Hello,
I have a question, If I have 10 Bam files (RNAseq) from 10 different patients and I want to see the different expression of few genes, what is the best tool to use after counting the reads?

Thank you

GenoMax · 03-05-2019, 08:37 AM

featureCounts part of Subread package. You can feed all your BAM files at the same time to get a matrix of counts (rows as genes and columns as your samples).

NDUFB11 · 03-05-2019, 08:44 AM

Thank you genoMax for your answer,

can you tell me how this reads are normalized?

thanks

GenoMax · 03-05-2019, 08:49 AM

You will get raw counts from featureCounts. You will need to use DESeq2/edgeR etc to actually do normalization and analysis.

NDUFB11 · 03-05-2019, 08:59 AM

but if I use Deseq2 it requires biological replicates, which I don't have it in my case.

How can I create the data frame and the condition with DESq2 in this case?

thank you

GenoMax · 03-05-2019, 09:54 AM

You can use DESeq2 without replicates. It is not recommended. Your analysis would not have any statistical significance.

Hopefully you have some other condition that you could test on?

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03-05-2019, 07:50 AM	#1
NDUFB11 Member Location: Italy Join Date: Jul 2017 Posts: 34	RNAseq from patients Hello, I have a question, If I have 10 Bam files (RNAseq) from 10 different patients and I want to see the different expression of few genes, what is the best tool to use after counting the reads? Thank you

03-05-2019, 08:59 AM	#5
NDUFB11 Member Location: Italy Join Date: Jul 2017 Posts: 34	but if I use Deseq2 it requires biological replicates, which I don't have it in my case. How can I create the data frame and the condition with DESq2 in this case? thank you Last edited by NDUFB11; 03-05-2019 at 09:03 AM.

03-05-2019, 08:37 AM	#2
GenoMax Senior Member Location: East Coast USA Join Date: Feb 2008 Posts: 7,087	featureCounts part of Subread package. You can feed all your BAM files at the same time to get a matrix of counts (rows as genes and columns as your samples).

03-05-2019, 08:44 AM	#3
NDUFB11 Member Location: Italy Join Date: Jul 2017 Posts: 34	Thank you genoMax for your answer, can you tell me how this reads are normalized? thanks

03-05-2019, 08:49 AM	#4
GenoMax Senior Member Location: East Coast USA Join Date: Feb 2008 Posts: 7,087	You will get raw counts from featureCounts. You will need to use DESeq2/edgeR etc to actually do normalization and analysis.

03-05-2019, 09:54 AM	#6
GenoMax Senior Member Location: East Coast USA Join Date: Feb 2008 Posts: 7,087	You can use DESeq2 without replicates. It is not recommended. Your analysis would not have any statistical significance. Hopefully you have some other condition that you could test on?